Clinical Research and Public Health
Abstract
BACKGROUND. Approaches to achieving antiretroviral therapy (ART)-free remission from HIV-1 must consider that people over 50 years now comprise the majority of people with HIV (PWH) on ART in various regions, including the U.S. METHODS. We report a double-blind, randomized trial in which PWH on ART, aged 21-60 years, received modified vaccinia Ankara (MVA)-vectored vaccines, MVA.tHIVconsv3 (M3) and MVA.tHIVconsv4 (M4), either alone or in combination (n=7/group) or saline placebo (n=3). M3 and M4 contain complementary HIVconsvX immunogens that each span the same regions in HIV-1 Gag and Pol but differ at approximately 8% at the amino acid level. RESULTS. M3, M4, and M3M4 regimens were well tolerated and all significantly increased both the frequency (peak median increase ~3-fold) and breadth of the HIVconsvX-specific T-cell response while redirecting T cells to target conserved regions in HIV-1 for up to 10 weeks post-vaccination. We also demonstrated that vaccination increased frequencies of T-cells targeting participant autologous HIV-1 sequences. Vaccination mostly expanded pre-existing HIV-1-specific T cells, did not impact CD4 T-cell activation, low-level viremia, or integrated HIV-1 provirus. Linear regression indicated that age was independently and negatively associated with the change in T-cell frequency at 1-, 2- and 10-weeks after vaccination (~1.41-fold decrease per 10 years older). After adjusting for age, years on ART was positively associated with HIVconsvX-specific T-cell frequencies at 1- and 2-weeks following vaccination. CONCLUSION. In PWH receiving ART, MVA.HIVconsvX vaccines significantly increased T cells targeting conserved regions of HIV-1. Novel strategies may be required to enhance anti-HIV-1 immunity in older adults. TRIAL REGISTRATION. NCT03844386.
Authors
Cynthia L. Gay, Yinyan Xu, Ann Marie K. Weideman, Fiona R. Shaw, JoAnn D. Kuruc, Shayla Z. Conrad, Sofia A. Mariano, Shahryar Samir, Sallay Kallon, Alexis T. Sponaugle, Joanna A. Warren, Genevieve T. Clutton, Maria Abad-Fernandez, Carolina Kapper, Alex B. Bradley, Caroline E. Baker, Susan M. Pedersen, Matthew Moeser, Lauren Burke, Edmund G.T. Wee, Alison Crook, Gregory M. Laird, Joshua C. Cyktor, John W. Mellors, Shuntai Zhou, Lawrence Fox, Joe J. Eron, David M. Margolis, Michael G. Hudgens, Tomáš Hanke, Nilu Goonetilleke
Abstract
BACKGROUND. Primary therapy for high-risk bladder cancer (BCa) is repeated instillations of the tuberculosis vaccine Bacillus Calmette-Guerin (BCG). Although BCG reduces the risk of recurrence by more than half, the mechanisms underlying its immune-activating effects remain unknown. Our objective was to investigate how the immune response differs between BCG responders and non-responders and to compare systemic and local immune responses. METHODS. We performed single-cell RNA sequencing (scRNA-seq) of isolated immune cells adjacent to high-risk bladders in BCG responders and non-responders before and after BCG. We also compared concurrent scRNA-seq profiles of circulating immune cell populations with those of bladder immune cells. RESULTS. We identify an increase in Th17-like Th1 cells in BCG responders, characterized by greater expression of pro-inflammatory cytokines. Alternatively, non-responders show increased CD8+ T-cell exhaustion and T regulatory cells. We identify that the primary mechanism driving divergent T-cell activity is altered polarization and immunosuppressive signaling with myeloid cells. Using a machine-learning-based approach, we identify that Th17-like Th1 cytokines, such as IL-17, IL-21, and IL-26, are predictive of response, which is subsequently validated in a separate BCG-treated BCa cohort. CONCLUSION. Together, these findings suggest that dynamic regulation of myeloid-T cell interactions can be critical for outcomes of BCG treated bladder cancer.
Authors
Ryan J. Brown, Mairah T. Khan, Andrew J. Houston, Hongshen Niu, Joseph R. Podojil, Bonnie Choy, Weiguo Cui, Joshua James Meeks
Abstract
BACKGROUND. Anti-nephrin autoantibodies have emerged as a putative pathogenic driver in a subset of patients with podocytopathies, including those with post-transplant disease recurrence. METHODS. We measured anti-nephrin autoantibodies in a cohort of 65 patients with podocytopathy associated with steroid-sensitive nephrotic syndrome (n = 39) and steroid-resistant nephrotic syndrome (n = 26), and in 34 patients with post-transplant podocytopathy recurrence. Fourteen patients with membranous nephropathy and 20 healthy volunteers served as controls. ELISA and immunoprecipitation assays were performed to detect anti-nephrin IgG using two different recombinant human nephrin proteins. Immunofluorescence analysis was performed to assess the deposition of IgG and their colocalization with nephrin in renal biopsies. RESULTS. When using murine antigen-based ELISA, the highest positivity was found in healthy volunteers (55%), correlating with levels of circulating natural anti-α-galactose-α-1,3-galactose antibodies. This cross-reactivity was abrogated with recombinant human nephrin expressed in human cells. In this setting, very low prevalence (<5%) of anti-nephrin antibody-positive patients was found in steroid-sensitive and steroid-resistant nephrotic syndrome cohorts and in patients with post-transplant disease recurrence. These frequencies were comparable to healthy volunteers. Using confocal and super-resolution microscopy, only trace amounts of IgM, but no IgG, were found in the glomeruli of analyzed biopsies, which did not colocalize with nephrin. CONCLUSIONS. With the methodology presented here, anti-nephrin reactivity was extremely rare and occurred at comparably low frequencies in healthy controls, native-kidney podocytopathies, and post-transplant disease recurrence. This suggests that these autoantibodies are not inherently disease-specific and may not serve as a broad biomarker across podocytopathies. TRIAL REGISTRATION. ClinicalTrials.gov NCT06334692. FUNDING. Private donation.
Authors
Francesco Pecoraro, Luca Perico, Federica Casiraghi, Paola Rizzo, Matias Trillini, Andrea Angeletti, Manuel Alfredo Podestà, Xhuliana Kajana, Agnese Spennacchio, Marta Todeschini, Marilena Mister, Giuseppe Castellano, Ariela Benigni, Giuseppe Remuzzi
Abstract
BACKGROUND. Elevated lipoprotein(a) [Lp(a)] is associated with a higher risk of atherosclerotic cardiovascular disease (ASCVD). Although Lp(a) is a genetically determined risk factor, the plasma proteomic features associated with Lp(a) and whether they provide information about ASCVD risk beyond Lp(a) concentration are not well characterized. OBJECTIVE. We sought to identify plasma proteomic features associated with Lp(a) concentration and to evaluate whether an Lp(a)-associated proteomic signature is associated with ASCVD phenotypes in young, healthy adults. METHODS. In the Coronary Artery Risk Development in Young Adults (CARDIA) study, we measured Year 7 Lp(a) and 184 cardiovascular proteins using the Olink proximity extension assay in 3,920 participants without prior coronary heart disease. Lp(a)-associated proteomic signatures were derived using LASSO regression in a split-sample design and tested for association with coronary artery calcification (CAC), incident CHD, and hs-CRP over 27 years of follow-up. External replication was performed in the UK Biobank (n=37,996). RESULTS. Lp(a) was associated with CAC (OR 1.23 [1.13-1.34]; p<0.0001) and incident CHD (HR 1.23 [1.07-1.41]; p=0.004). Lp(a) correlated with proteomic features reflecting immune activation, coagulation, and vascular dysfunction. A quantitative Lp(a) proteomic score was independently associated with incident CAC (standardized beta = 0.40, p<0.0001) and hs-CRP (standardized beta = 0.11, p = 0.00015) after adjustment for Lp(a) concentration. In the UK Biobank, a recalibrated Lp(a)-associated proteomic score was associated with CRP, incident CHD, and all-cause mortality. CONCLUSIONS. In young adults, Lp(a) is associated with distinct proteomic features that independently predict ASCVD phenotypes beyond Lp(a) concentration, generating hypotheses regarding biological pathways linked to Lp(a)-related cardiovascular risk.
Authors
Sascha N. Goonewardena, Shanshan Yao, Tomasz Jurga, Lanyue Zhang, Donald Lloyd-Jones, Dilna Damodaran, Bharat Thyagarajan, David R. Jacobs Jr, Supriya Shore, Eric J. Brandt, Clary Clish, Kahraman Tanriverdi, Jane E. Freedman, Chirag J. Patel, Mark A. Sarzynski, Brian T. Emmer, John T. Wilkins, Ron Do, Vera Bittner, Ravi Shah, Marios K. Georgakis, Robert S. Rosenson, Venkatesh Murthy
Abstract
BACKGROUND. Current diagnosis and surveillance of bladder cancer relies on cystoscopy which is invasive and user dependent. The urine mRNA panel, uRNAp, measures expression of 3 genes for identification of bladder cancer. Here we report validation of uRNAp for patients undergoing initial work-up for suspected bladder cancer and surveillance for bladder cancer. METHODS. Urine specimens were prospectively collected prior to cystoscopy at two health care systems from patients without (detection cohort) or with (surveillance cohort) a history of bladder cancer. RNA was isolated from urine sediment for RT-qPCR to determine ROBO1, CRH, and IGF2 expression and calculate the uRNAp bladder cancer probability score. RESULTS. In the detection cohort, 547 samples were collected from 529 patients. There were 123 new diagnoses of bladder cancer in the detection cohort and uRNAp demonstrated 98% sensitivity and 51% specificity for identification of bladder cancer. In the surveillance cohort, 1543 samples were collected from 447 patients with 286 recurrences. uRNAp demonstrated 94% overall sensitivity with 43% specificity and 99% sensitivity for high-grade recurrence. The receiver operating characteristic area under the curve was 0.92 in the detection and 0.81 in the surveillance cohort. uRNAp scores significantly increased with tumor size and grade. CONCLUSIONS. Prospective validation of uRNAp demonstrated a strong potential clinical utility as a non-invasive adjunct to cystoscopy for management of bladder cancer. uRNAp may be a useful triage tool to defer or expedite cystoscopy for patients undergoing detection or surveillance of bladder cancer. FUNDING. Department of Veterans Affairs BLR&D Merit Review I01 BX004962 to JCL.
Authors
Kathleen E. Mach, Zachary Kornberg, Eugene Shkolyar, Jin Long, Timothy J. Lee, Vinh La, Ihna Yoo, Gabriela Rodriguez, Alan E. Thong, Kris B. Prado, Jay B. Shah, John T. Leppert, Eila C. Skinner, Joseph C. Liao
Abstract
Background. Functional B cell responses for both prevention and control of hepatitis B virus (HBV) infection remain poorly understood, including in the context of HBV/HIV co-infection. Methods. Here, we employed high-dimensional single cell analysis to assess global and hepatitis B surface antigen (HBsAg)-specific B cells in a longitudinal cohort of incident HBV from the Multicenter Aids Cohort Study (MACS), with a subset of the cohort living with HIV-1. Results. We observed that prior HIV infection has negative consequences for B cell function in early post-acute HBV infection, including increased frequencies of atypical memory (AtM) B cells and regulatory B cells (Bregs), expression of the activation marker CD86 on multiple B cell subsets in chronic HBV (CHB), and restricted expansion of HBsAg-specific B cells. In contrast, in HBV mono-infection, we observed no changes in the global B cell population from prior to infection and robust expansion of HBsAg-specific B cells. These expanded antigen-specific B cells resembled class-switched intermediate and resting memory (IM and RM) B cells, with activation phenotypes that may contribute to ongoing HBV control. Conclusion. HIV infection has a significant impact on B cell responses to subsequent HBV infection that may promote development of CHB in HBV/HIV co-infection. Funding. National Institute of Allergy and Infectious Diseases, Bill & Melinda Gates Foundation.
Authors
Katherine Cascino, Thomas Liechti, Eric C. Seaberg, Kathleen E. Stevens, Steven M. Wolinsky, Mallory D. Witt, Robbie B. Mailliard, Mario Roederer, Justin Bailey, Chloe L. Thio, Andrea L. Cox
Abstract
BACKGROUND Sepsis encompasses considerable biological and clinical heterogeneity. Previously, 2 phenotypes (“hyperinflammatory” and “hypoinflammatory”) have been consistently identified within sepsis via latent class analysis. These phenotypes differ in their biological features, clinical outcomes, and therapeutic responses to interventions. Prior studies of sepsis heterogeneity have focused primarily on the host response. Here, we investigate the potential influence of the causative pathogen on sepsis heterogeneity and pathobiology.METHODS We performed a retrospective observational analysis of 8,280 critically ill patients with sepsis to identify associations between pathogen characteristics and the hyperinflammatory and hypoinflammatory patient phenotypes. We also performed controlled murine and swine modeling of sepsis and lung injury and a secondary analysis of 449 patients enrolled in the EUPHRATES randomized controlled trial.RESULTS Pathogen characteristics (pathogen identity, burden, virulence, and anatomic site of infection) were strongly and independently associated with the previously reported phenotypes. In a cohort of critically ill patients with sepsis, infection with gram-negative pathogens, primarily Enterobacterales spp. (e.g., Escherichia coli, Klebsiella pneumoniae), was strongly associated with the hyperinflammatory phenotype. The hyperinflammatory phenotype was also independently associated with increased pathogen burden, virulence, and initial anatomic site of infection. In controlled murine and swine modeling, both the identity and burden of the pathogen provoked key biological features of the hyperinflammatory phenotype. Among patients with sepsis, the prognostic value of lactate clearance varied substantially by phenotype. In a secondary analysis of a randomized trial of polymyxin B hemoadsorption (which removes circulating endotoxin), hypoinflammatory patients experienced worse survival.CONCLUSIONS Our results demonstrate the central importance of pathogen features in the clinical and biological heterogeneity of sepsis. Future studies of sepsis pathobiology and heterogeneity should expand their scope beyond the host response, as understanding pathogen-host interactions will be crucial in the development of precision therapeutic strategies to improve patient outcomes.TRIAL REGISTRATION EUPHRATES trial NCT01046669.FUNDING 5P30AG024824, IK2CX002766, R01HL144599, K24HL159247, R01HL158626, R01HL173531, R35GM142992, R35GM145330, R35GM136312, K23HL166880, R35HL140026.
Authors
Rishi Chanderraj, Brian Bartek, Kathleen A. Stringer, Mohamad H. Tiba, Michael W. Sjoding, Ying He, Mark Nuppnau, Kale S. Bongers, Mark D. Adame, Sunny S. Lou, V. Eric Kerschberger, Matthew M. Churpek, Carolyn S. Calfee, Sandhya Tripathi, Debra M. Foster, John A. Kellum, Robert P. Dickson, Pratik Sinha
Abstract
BACKGROUND Gut microbes and their metabolites contribute to the host circulating metabolome and exhibit diurnal variation influenced by sleep-wake cycles and meal timing. Sleep deprivation alters the rhythmic circulating metabolome, but its impact on microbial metabolites remains unclear. We tested whether 24-hour circulating metabolite profiles, including those of microbial origin, differ under normal (habitual) versus short-term restricted sleep.METHODS In a randomized crossover design, 9 healthy adults completed 2 in-lab 24-hour blood sampling sessions (q120): one following 3 nights of normal sleep (8.5 hours/night), the other following 3 nights of sleep restriction (4.5 hours/night). Meal timing and caloric intake were held constant. Serum metabolites were characterized using untargeted reverse-phase liquid chromatography–mass spectrometry and rhythmicity was assessed using empirical JTK_CYCLE analysis.RESULTS We identified 90 metabolites, including 14 of microbial origin or derived from host metabolism of microbial products, e.g., butyrate and tryptophan derivatives. Sleep restriction significantly altered serum metabolite composition compared with normal sleep. While many compounds maintained rhythmicity across conditions, sleep restriction disrupted rhythms of several key compounds, including microbe-derived metabolites. Notably, butyrate and indole-3–propionic acid lost rhythmicity, whereas new rhythms emerged in the tryptophan catabolite, kynurenine, and lipid metabolism intermediates.CONCLUSION We provide evidence that microbial metabolites are detectable in human blood and exhibit sleep-dependent rhythmicity. Sleep restriction alters diurnal circulating microbial and host-derived metabolite rhythms even under constant meal timing, composition, and calories. These findings support links between host sleep patterns and gut microbial metabolism and suggest microbial metabolites as potential biomarkers or mediators of sleep loss–associated health risks.TRIAL REGISTRATION NCT00989976.FUNDING NIH/NCRR KL2RR025000; R56DK102872-01A1, P30DK020595; P30DK042086; K01DK111785; F31DK122714; DOD W81XWH-07-2-0071.
Authors
Vanessa A. Leone, Katya Frazier, Manpreet Kaur, Evan A. Chrisler, Ashley M. Sidebottom, Ethan Tai, ViLinh Tran, Shuzhao Li, Eugene B. Chang, Dean P. Jones, Eve Van Cauter, Erin C. Hanlon
Abstract
BACKGROUND. Immune checkpoint inhibitors (ICIs) targeting the programmed cell death-1 axis have revolutionized metastatic non–small cell lung cancer (mNSCLC) treatment. However, disease progression remains a concern, and the role of the complex tumor microenvironment (TME) in treatment failure is not fully understood. METHODS. In this biomarker study involving 103 patients with mNSCLC—including 81 patients who received ICI treatment—we evaluated the association between heterogeneous immune cell subsets and ICI efficacy through single-cell spatial profiling of pretreatment tumor tissue, using a 29-marker multiplex immunohistochemistry platform built for in-depth dissection of the TME. RESULTS. Among various types of intratumoral lymphocytes including T-helper 1 cells, regulatory T cells, and natural killer cells, only CD8+ T cells (TILs) were associated with ICI efficacy. Computational tissue segmentation underscored the importance of direct physical interactions between CD8+ TILs and cancer cells for ICI efficacy. TIL phenotyping identified CD39/CD103/Ki-67 positivity as a hallmark of exhausted yet functional tumor-reactive CD8+ TILs. Immunosuppressive tumor-associated macrophages (TAMs) and cancer-associated fibroblasts were independent unfavorable adversaries. High CD73 expression on cancer cells was suggested to confer tolerance to ICI in EGFR/ALK-oncogene+ NSCLC, potentially through M2-TAM accumulation and aberrant angiogenesis. CONCLUSION. Our study delineates the clinical relevance of heterogeneous immune cell subsets in ICI-treated mNSCLC, aiding the development of targeted therapeutic strategies. TRIAL REGISTRATION. Not applicable because this is a retrospective study. FUNDING. Osaka Cancer Society, KANAE Foundation for the Promotion of Medical Science, SGH Foundation, and YOKOYAMA Foundation for Clinical Pharmacology.
Authors
Kohsuke Isomoto, Koji Haratani, Takahiro Tsujikawa, Shuta Tomida, Yusuke Makutani, Masayuki Takeda, Kimio Yonesaka, Kaoru Tanaka, Tsutomu Iwasa, Kazuko Sakai, Kazuto Nishio, Akihiko Ito, Kazuhiko Nakagawa, Hidetoshi Hayashi
Abstract
BACKGROUND. Genetically engineered porcine livers are being developed as a bridge therapy for acute liver failure, providing detoxification and restoration of hepatic protein synthesis. Severe xenograft-associated severe thrombocytopenia remains a major limitation, and human mechanistic data are scarce. METHODS. Platelet kinetics were characterized in three human decedents undergoing extracorporeal cross-circulation with transgenic porcine livers. Platelet counts, transfusion requirements, and clearance patterns were assessed to distinguish consumption from marrow suppression or hypersplenism. Antibody- and complement-directed inhibitors were administered to test immune-mediated mechanisms. Mechanistic studies focused on porcine von Willebrand factor (pVWF)–dependent platelet activation, including ex vivo blockade with the anti-VWF nanobody caplacizumab, a vWF-directed antibody fragment that prevents vWF–platelet binding. A fourth decedent received caplacizumab during porcine liver perfusion. RESULTS. In all three initial cases, 80%–90% of circulating and transfused platelets were rapidly cleared, a pattern inconsistent with marrow suppression or hypersplenism. Antibody and complement inhibition failed to ameliorate thrombocytopenia. Recipient plasma induced robust pVWF-mediated platelet activation analogous to human Type IIb von Willebrand disease, which was completely abrogated ex vivo by caplacizumab. In a fourth decedent treated with caplacizumab, aberrant platelet activation was prevented, though full hematologic recovery was limited by pre-existing disseminated intravascular coagulation (DIC). CONCLUSIONS. Early thrombocytopenia during porcine liver xenotransplantation appears to be primarily driven by pVWF-mediated platelet activation rather than by classical immune or splenic mechanisms. Targeted VWF blockade with agents such as caplacizumab may mitigate platelet loss and improve the safety profile of extracorporeal porcine liver support in acute liver failure.
Authors
Liang Zhao, Sokratis A. Apostolidis, Aae Suzuki, Amrita Sarkar, Qian Guo, Felix Li, Alex Sagar, John I. Fallon, Mohamed A. Elzawahry, Syed Hussain Abbas, Leanne Lanieri, Kristen Getchell, Susan C. Low, Kim M. Olthoff, Emma E. Furth, Brendan J. Keating, Peter Friend, Mortimer Poncz, Abraham Shaked, Charles S. Abrams
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